Inhibition of Neuronal Nitric Oxide Synthase by 4-Amino Pteridine Derivatives:  Structure−Activity Relationship of Antagonists of (6R)-5,6,7,8-Tetrahydrobiopterin Cofactor

Abstract

The family of nitric oxide synthases (NOS) catalyzes the conversion of l-arginine to l-citrulline and nitric oxide (NO), an important cellular messenger molecule which has been implicated in the pathophysiology of septic shock and inflammatory and neurodegenerative disease states. NOS can be maximally activated by the ubiquitous cofactor, (6R)-5,6,7,8-tetrahydrobiopterin (H4Bip), and antagonists of H4Bip may be of therapeutic importance to inhibit pathologically high NO formation. The 4-amino substituted analogue of H4Bip was reported to be a potent NOS inhibitor. Therefore, we developed a series of novel 4-amino pteridine derivatives, anti-pterins, to pharmacologically target the neuronal isoform of nitric oxide synthase (NOS-I). To functionally characterize the pterin/anti-pterin interaction and establish a structure−activity relationship (SAR), we systematically altered the substituents in the 2-, 4-, 5-, 6-, and 7-position of the pteridine nucleus. Varying the substitution pattern in the 2-, 5-, and 7-position resulted in no significant inhibitory effect on enzyme activity. In contrast, bulky substituents in the 6-position, such as phenyl, markedly increased the inhibitory potency of the reduced 4-amino-5,6,7,8-tetrahydropteridines, possibly as a consequence of hydrophobic interactions within NOS-I. However, this was not the case for the aromatic 4-amino pteridines. Interestingly, chemical modification of the 4-amino substituent by dialkyl/diaralkylation together with 6-arylation of the aromatic 2,4-diamino pteridine resulted in potent and efficacious inhibitors of NOS-I, suggesting possible hydrophilic and hydrophobic interactions within NOS-I. This SAR agrees with (a) the recently published crystal structure of the oxygenase domain of the inducible NOS isoform (NOS-II) and (b) the comparative molecular field analysis of selected NOS-I inhibitors, which resulted in a 3D-QSAR model of the pterin binding site interactions. Further optimization should be possible when the full length structure of NOS-I becomes available.

Synthesis of New 2‐Phenyladenines and 2‐Phenylpteridines and Biological Evaluation as Adenosine Receptor Ligands

“…The reaction of benzamidine chloride with a slight excess of silver salt of isonitrosomalononitrile gave 4,6-diamino-5-nitroso-2-phenylpyrimidine 16 as described in the literature [29,30]. Compound 16 was treated with sodium hydrosulphite to give, by reduction of the nitroso group, 4,5,6-triamino-2-phenylpyrimidine 17 [30,31]. Cyclisation of 17 with aqueous glyoxal furnished 18 [31] which reacted with the suitable isocyanates to give the 1-substituted phenyl-3-(2-phenyl-pteridin-4-yl)-ureas 19 and 20.…”

Computational Development of Selective nNOS Inhibitors: Binding Modes and Pharmacokinetic Considerations

“…Matter et al developed a series of 65 4-amino pteridine and 4-oxo pteridines, and examined their inhibitory effects on recombinantly produced NOS protein [35][36][37]. In the absence, at the time, of any experimentally derived protein crystal structure for nNOS, Matter et al made use of homology modelling to rationally develop these 65 inhibitors.…”

Biology and chemistry of the inhibition of nitric oxide synthases by pteridine‐derivatives as therapeutic agents

“…Recently a series of pterin-based inhibitors as NOS-I inhibitors without effect on phenylalanine hydroxylase was discovered. 98,106,107 This led to the design and synthesis of structural analogs as NOS inhibitors based on the 4-oxo pteridine scaffold. Systematic variations, docking and X-ray structure analysis of inhibitors in the homologue NOS-III oxygenase domain established the structure-activity relationship and resulted in the discovery of potent and effective NOS-I inhibitors.…”

“…Systematic variations at the 2-, 4-, 5-, 6-, and 7-position for these 4-amino pteridine derivatives led to develop a structure-activity relationship for NOS-I. 98,107 No significant inhibition on NOS-I enzyme activity was observed upon substitution of positions 2, 5, and 7, whereas bulkier substituents in position 6, such as phenyl, led to significantly increased affinity of reduced 4-amino-5,6,7,8-tetrahydropteridines (Fig. 5, upper), because of favorable hydrophobic interactions to NOS-I.…”